Aims The aim of this PhD research project was to investigate the application of dental pulp stem cells (DPSC) as a treatment for traumatic and degenerative eye diseases. The accuracy and reliability of counting retinal ganglion cells (RGC) in radial retinal section was also assessed. Methods Numbers of RGC in radial retinal sections were compared to numbers in retinal wholemounts. DPSC were cultured with RGC and survival and neuritogenesis were quantified. DPSC were also transplanted intravitreally into rat models of optic neuropathy (optic nerve crush) and glaucoma and surviving RGC and regenerated axons were quantified in radial retinal sections. Results Quantifying RGC in radial retinal sections was as reliable and accurate as the current gold standard Thus, retinal wholemounts with Brn3a proved to be the most reliable marker for RGC. DPSC protected RGC from optic nerve crush-/glaucoma-induced death, promoting significant regeneration of RGC axons in the former and preserving visual function (as measured by electroretinography) in the latter. The mechanism of action, as determined in vitro, appeared to be through the secretion of multiple neurotrophic factors (NTF). Conclusions In conclusion, DPSC is a potent cell therapy in the treatment of traumatic and degenerative eye disease.

No description available.

Differential growth was demonstrated between facial features and within some facial features. In particular, the columella, nostrils and philtrum did not grow significantly after surgery, although this would be considered normal in the age group studied. Facial growth in children with UCL and UCLP was independent of the head and body growth. The presence of a cleft of the secondary palate accentuated the amount of soft tissue disruption by the cleft in the lip and nose, but not the pattern of disruption. Primary lip / nose repair had no detrimental effect on the early growth and development of the facial features. Likewise, palate repair had no discernible effect on facial soft tissue growth at age 2 years. Primary lip /nose repair had a beneficial effect on facial morphology in terms of reducing asymmetry and was most successful in the improving philtrum and nasal base symmetry, less successful in improving the nasal rim asymmetry. A possible early beneficial effect of cleft repair remote from the surgery site was noted in the reduction of upper face asymmetry in the first year of life. Residual asymmetry in the facial features did not change by age 2 years, despite increases in size with growth. Facial morphology outcomes for UCL and UCLP children in this study was generally similar at 2 years of age, despite marked differences in pre-operative facial form. However, nasal base asymmetry, upper face asymmetry and residual nostril shape deformity were significantly greater in UCLP children at 2 years of age, than in UCL children. These shape differences were not detectable by measurement of facial dimensions alone.

Background: Changes in clinical management; advances in non-invasive three-dimensional imaging; developments in methods of shape analysis. Aim: To assess three-dimensional dentofacial deformity with a view to early appraisal of primary surgical outcome. Results: Significant differences in upper lip morphology were found between the cleft children and their unaffected peers; nasal asymmetry that became more obvious in function was noted in cleft children; the maxillary dental arches of the children with repaired cleft palate were shallow, short and narrow; and the dental arch, deformity and the facial soft tissue deformity were unrelated. Contributions to the field: It has been shown that deviation from normal could be detected as young as 3 years of age using computerised stereophotogrammetry; preliminary, objective, three-dimensional analysis of facial function has been completed in young children; the accuracy of three-dimensional CT scanning of dentate study models and the time cost of data collection were quantified; and this study has produced a body of three-dimensional data that can test and support analytical advances.

Having established the antiquity of dentistry and its relationship to medicine, this thesis examines its development in Scotland from the unique origins in Glasgow of the Royal Faculty of Physicians and Surgeons and the Royal College of Surgeons, Edinburgh, to the legitimization of the profession by State intervention in 1921. Although an LDS qualification was available in 1860 from the Royal College of Surgeons, England, the introduction of the Dentists Act of 1878 and the establishment of a Register, brought about through the efforts of the Reform Movement, did not eradicate the practice of dentistry by the unregistered. Their numbers continued to multiply as a result of urban migration, socio-economic changes and industrialisation. Changes in the dietary habits of the Scottish people and in the methods of food production, contributed to an increase in the prevalence of dental caries as well as in the demand for dental treatment among all classes of the population, as epidemiological surveys and Government inquiries showed. The introduction of a school dental service resulted, when attention was focused on the poor dental standards of the recruits following the defeats of the British Forces in the Boer Wars. The resultant Government inquiry into the state of dentistry revealed the deplorable state of the nation's teeth and the inability of the Scottish working classes and the disadvantaged to afford dental treatment from qualified practitioners. In the aftermath of the 1914-18 War, public opinion was conducive to the idea of social welfare and public health thus paving the way for the legalization by the State of the dental profession by virtue of the Dentists Act of 1921.

An artificial oral environment was used to clinically simulate the dental wear process and surface friction between opposing maxillary and mandibular elements. Measurement of surface friction between occlusal surfaces of natural premolar teeth demonstrated large variation. High viscosity lubricants and surfactant containing lubricants were capable of reducing the surface friction. The enamel appeared otherwise insensitive to natural and artificial salivary fluids. Orientation of opposing enamel prisms was critical for reduction of interocclusal friction. A critical choice of prosthetic dental materials evaluted for wear included amalgam, porcelain, and composite opposed by a natural third molar palatal cusp. The wear of amalgam was slight (volume loss 0.0307 .0036 mm^3 and depth .072 .0017 mm at 300K masticatory cycles) and wear of the opposing enamel was not measurable. SEM demonstrated smearing between the amalgam and the enamel cusp, thus supporting an adhesive wear mechanism. The wear of porcelain (volume .165 .037 mm^3, depth .127 .02 mm at 300K masticatory cycles) demonstrated a ploughing effect on the surface supporting an abrasive wear mechanism. The composite wear was intermediate(volume .046 .007 mm^3, depth .059 .01 mm). Posterior composites are thought to wear by an abrasive as well as fatigue mechanism. The artificial mouth demonstrated a high correlation (0.98) with clinical studies.Wear of enamel opposed by enamel demonstrated a degree of variability. The physiologic occlusal orientation between enamel pairs appeared important. A finite element model of a natural and crowned premolar tooth was developed consisting of 586 triangular elements and 343 nodes using ANSYS and IFECS finite element analysis software. Validation of the model was achieved via a physical model of a natural premolar tooth with strain gages. A new full coverage restoration design employing layering of composite restorative materials was proposed and tested. The model predicted a problem of fatigue crack propagation through the restoration which was confirmed in preliminary clinical trials. This provided further evidence for the fatigue mechanism of wear in composite restorations. On the basis of the wear studies conducted, recommendations for restorative treatment design were presented.

Denture-associated stomatitis (DS), a frequent infection in denture-wearers (up to 60%), presents as areas of palatal inflammation and is normally associated with denture biofilms containing Candida albicans. However, the contribution of co-existing bacteria in these biofilms to the infection remains unclear. As current DS management strategies are primarily directed towards Candida, research demonstrating the impact of specific bacteria upon infection prognosis is important to improve treatment regimes. This research evaluated the in vitro impact of bacteria on Candida virulence, and compared bacterial microbiomes at specific oral sites in DS and non-DS patients to determine associations with infection. In vitro biofilm studies assessed expression of C. albicans virulence factors (morphological transformation, adhesins, hydrolytic enzymes) and their impact on pathogenesis in an infection model. In clinical studies, microbiological samples were obtained from the tongue, palate and denture-fitting surface of 19 denture-wearing patients (DS n=8, non-DS n=11). The presence of Candida was ascertained by PCR. Bacterial DNA was extracted and subjected to next generation sequencing using bacterial 16S rRNA gene targets, and differences in the bacterial microbiomes determined. Certain bacterial species in acrylic biofilms significantly (P < 0.05) increased the expression of C. albicans virulence factors, and subsequently, enhanced tissue damage in model systems. Candida was detected in clinical samples of 14 patients (DS n=6, non-DS n=8). Metataxonomic analyses revealed differences in relative abundance of bacterial species, but no significant differences in the bacterial microbiomes of the denture-fitting surface and palate between DS and non-DS patients. Importantly, a significant (P=0.007) increase in the number of bacterial species was evident for the tongue microbiome of non-DS patients. The in vitro modulating capacity of bacteria toward Candida virulence, and the observed species-level differences in bacteria between DS and non-DS patients highlight the need for consideration of the bacterial composition of oral biofilms in the pathogenesis of DS.

Candida infections of humans are an increasingly prevalent problem. Most candidal infections are superficial, occurring on mucosal surfaces, however, systemic infection can arise in immunocompromised individuals and these are life threatening. Candida is recognised by host immune cells through pattern recognition receptors e.g. dectin-1. The immune cells then produce cytokines to drive adaptive immune responses. The type of T-helper (Th) cell response that occurs is an important factor in whether candidal colonisation or clearance occurs. The overall focus of this research was to employ in vivo and in vitro studies to assess the nature of the immune response to Candida albicans. Increased pro-inflammatory Th17 and Th1 responses were evident in denture stomatitis (DS) patients based on cytokine profiles. In chronic hyperplastic candidosis (CHC) tissues significantly higher levels of CD4+, IL-12A+, IL-17A+ and EBi3+ positive cells were detected by immunohistochemistry (IHC) compared to control tissues. This finding was indicative of an increased Th17 response in CHC. IHC detection of individual cytokine subunits in cells was more difficult to interpret, but suggested both IL-35 and IL-12 cytokines were present, indicating Treg and Th1 cell responses, respectively. Challenge of a human monocyte cell line and human peripheral blood mononuclear cells (PBMCs) with C. albicans resulted in IL-23 cytokine expression indicative of a Th17 response. Interestingly, lipopolysaccharide (LPS) was important in enhancing the ability of dendritic cells to recognise and phagocytose C. albicans via dectin-1. The recall response from PBMCs stimulated with C. albicans resulted in a significant Th17 recall response. Extrapolation of these findings to the host interaction with Candida requires additional clinical studies and assessment of other forms of superficial mucosal candidosis. This research does however indicate that host recognition of C. albicans leads to a predominantly pro-inflammatory Th17 response.

Recurrence and persistence of microbial infection is one of the main reasons for the revision of dental restorations in the clinic. Failure to control the pathogenic microbiota leads to the formation of caries, which in turn may lead to irreversible pulpal damage,resulting in the need for root canal (endodontic) therapy. Secondary endodontic infections can spread to the surrounding oral tissues and beyond, leaving the patient vulnerable to systemic infection. This project aimed to develop a novel, injectable hydrogel containing antimicrobial liposomes for use as an intracanal medicament in order to reduce the incidence of secondary endodontic infections. Triclosan, a broad-spectrum, hydrophobic antimicrobial drug, was shown to have a bacteriostatic and bactericidal effect against two oral pathogens, Enterococcus faecalis and Streptococcus anginosus, which were grown planktonically and as a single-species biofilm. The bacteriostatic effect was also seen when triclosan was encapsulated in multilamellar vesicles (MLVs) and small unilamellar vesicles (SUVs) of phosphatidylcholine:cholesterol (PC:C) liposomes. Antimicrobial efficacy was associated with a high drug:lipid ratio in the liposomes. The liposomes were incorporated into a methyl cellulose (MC) solution, and the rheological properties were measured. MC was a sheer-thinning, viscous solution at ambient temperature and formed a hydrogel as the temperature was increased above 30 C. These properties were unaffected by the addition of liposomal MLVs or SUVs. The hydrogels containing triclosan-loaded liposomes had an antimicrobial effect when incubated in contact with suspensions of E. faecalis or S. anginosus, but MC containing triclosan only did not. A release assay showed the release of triclosan from MC loaded with triclosan liposomes, which was not seen when triclosan was incorporated into MC alone. The liposomal hydrogel was injected into endodontically prepared extracted human teeth that had been inoculated with E. faecalis suspension to mimic endodontic infection. After 24 h treatment, histological analysis showed that triclosan solution,triclosan liposomes and triclosan liposomes in MC prevented the formation of a biofilm on the intraroot surface, which was observed in controls that underwent no treatment or treatment with MC hydrogel only. The results of this work indicated that triclosan liposomes have potential to prevent secondary endodontic infections and may be loaded into a hydrogel suitable for injection into the root canal and subsequent gelation upon thermoequilibriation with the oral cavity. Triclosan release from this hydrogel may facilitate the prevention of bacterial colonisation of the root canal by E. faecalis, which has high prevalance in secondary endodontic infections.

Background: Oral squamous cell carcinoma (OSCC) is the 15th most common cancer worldwide but has poor five year survival (50%). Late stage presentation and limitations of early diagnostic techniques are persistent clinical problems. Sixty percent of patients present with advanced stage disease and with the attendant increase in mortality, morbidity and risk of recurrent disease it is particularly burdensome for both patients and health economies. Early diagnosis and treatment of OSCC improves prognosis. There is an opportunity to diagnose OSCC early in patients with oral epithelial dysplasia however currently there is no way of accurately predicting which lesions will undergo malignant transformation. Aberrant methylation of tumour suppressor genes plays a significant role in the biology of early cancer and is detectable in both tumour and saliva. Saliva is a non-invasive method of longitudinal sampling and has potential as a tumour surrogate in disease surveillance programmes. This study aims to compare rates of methylation of a panel of genes in OSCC patients and a normal cohort to establish a threshold by which we could determine future disease testing in a dysplastic population. Methods: Saliva samples were collected from 219 individuals from three diagnostic groups: Normal (defined as no oral malignant or premalignant disease) n=97, OSCC n=62 and dysplasia n=60. For statistical analysis the dysplasia cohort was sub-divided into lesions of low and high risk of malignant transformation based on the histological diagnosis of the index lesion. DNA was extracted and bisulphite treated from 258 saliva samples before duplex quantitative methylation specific PCR (qMSP) assays were performed on all samples to detect the frequency of methylation in saliva of a panel of genes. The five target genes (ADAMTS9, CCNA1, CYGB, P16, TMEFF2) were selected using a candidate approach on the basis of tumour specificity from studies on tumour/normal matched tissue pairs. Clinicopathological data was correlated with the qMSP data and analysed using SPSS v.21 statistical software to look for associations with tumour and survival characteristics. Results: Only 3/97 individuals from the control normal cohort had saliva samples with detectable methylation above the analytical sensitivity of the P16 assay. Methylation of the remaining target genes (ADAMTS9, CCNA1, CYGB, TMEFF2) was not detected in normal saliva at levels above the analytical sensitivity of the qMSP assays. The most significant finding in this study was that methylation of four of the target genes (CCNA1, CYGB, P16, TMEFF2) in saliva, individually and when considered as a panel, was significantly associated with OSCC and as such could aid discrimination between malignant disease and normal saliva samples. Methylation of at least one gene in the panel was discovered in 29/67 of the binned OSCC saliva samples but only 3/97 of normal samples (Fisher’s exact p=0.001). Furthermore methylation of the gene panel is associated with high risk lesions when detected in saliva of patients with premalignant lesions (Fisher’s exact p=0.03). Conclusions: This exploratory data supports the utility of duplex qMSP as a detection method for methylation markers in saliva. The detection of methylation of this gene panel in saliva is significantly more associated with oral malignancy and high risk premalignant lesions than normal and low risk disease. This implies saliva may have merit as a surrogate tissue in an adjunctive role to clinical assessment and biopsy. The assays are specific but have limited sensitivity. However with further work, inclusive of additional genes, this methodology may identify predictive biomarkers that can be introduced into a trial surveillance of premalignant lesions.